Despite clear genetic evidence linking the beta-amyloid precursor protein (APP) to familial Alzheimer's disease (AD), the physiological functions of APP remain ill-defined. Our previous work suggests that APP plays a role in neddylation that activates Cullin ubiquitin ligase. In preliminary data, we show that APP modulated Cullin5 levels and mutant APPs caused excessive Cullin5 degradation. We found that downregulation of the endogenous APP in primary neurons resulted in less Cullin5 turnover. We further demonstrated that more APP knockout CA1 pyramidal cells have Cullin5 expression than the wild type counterparts. These data strongly suggest that APP modulates Cullin5 ligase activity under physiological conditions, and further suggest that increased APP gene dosage or APP mutations may impair Cullin5 ligase functions by increasing its turnover rate and leading to decreased clearance of signaling molecules and neuronal dysfunction. We also discovered that inactivation of the ubiquitin-activating enzyme E1 resulted in decreased levels of APP. Furthermore, E1 co-precipitated with APP in mouse brain homogenates. These data suggest that E1 may affect APP processing by binding to APP. In this application, we hypothesize that APP is downstream of the ubiquitin- activating enzyme E1 but upstream of the ubiquitin ligase Cullin5, thus playing an important physiological function in the clearance of signaling molecules such as tyrosine kinases. This hypothesis will be tested by four inter-related Specific Aims: Aim 1, Determine if APP regulates Cullin5 neddylation and function via BP1; Aim 2, Elucidate how APPswe destabilizes Cullin5; Aim 3, Determine if E1 activation increases amyloid genesis; Aim 4, Determine if Presenilin mediates Cullin5 turnover and activation. Successful completion of this project will establish APP as an important component of the ubiquitination pathway. Therefore, the results from testing this hypothesis will provide a new understanding of APP functions/dysfunctions, which may reveal targets for disease interventions and biomarker discoveries.